| 杨成杰,李英琦,黄星,等.毛蕊异黄酮通过 PI3K/Akt/mTOR信号通路对高糖作用下人视网膜色素上皮细胞自噬的影响[J].安徽医药,2026,30(7):1333-1338. |
| 毛蕊异黄酮通过 PI3K/Akt/mTOR信号通路对高糖作用下人视网膜色素上皮细胞自噬的影响 |
| Effect of calycosin on autophagy of human retinal pigment epithelial cells under high glucose through PI3K /Akt/mTOR signaling pathway |
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| DOI：10.3969/j.issn.1009-6469.2026.07.012 |
| 中文关键词: 黄芪 糖尿病视网膜病变 毛蕊异黄酮 人视网膜色素上皮细胞 高糖 磷脂酰肌醇 3-激酶 /蛋白激酶 B/哺乳动物雷帕霉素靶蛋白(PI3K/Akt/mTOR) 自噬 |
| 英文关键词: Astragalus membranaceus Diabetic retinopathy Calycosin Human retinal pigment epithelial cells High glu-cose Phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin(PI3K/Akt/mTOR) Autophagy |
| 基金项目:贵州省科技计划项目(黔科合基础 -ZK〔2023〕一般 396)；贵阳市科技计划项目(筑料合同〔2019〕9-1-8) |
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| 中文摘要: |
| 目的探讨毛蕊异黄酮(CAL)通过磷脂酰肌醇 3激酶 /蛋白激酶 B/哺乳动物雷帕霉素靶蛋白( PI3K/Akt/mTOR)信号通路对高糖作用下人视网膜色素上皮细胞( ARPE-19)自噬的影响。方法于 2022年 12月至 2023年 12月体外培养 ARPE-19细胞，分为对照组( 5.5 mmol/L葡萄糖)，高渗组( 50 mmol/L甘露醇)，高糖组( 50 mmol/L葡萄糖)，CAL低、中、高浓度组( 50 mmol/L葡萄糖 +25、50、75 μmol/L-CAL)筛选 CAL最佳浓度( 75 μmol/L)后，将细胞分为:正常对照组、高糖模型组、 CAL高浓度组( 50 mmol/L葡萄糖 +75 μmol/L-CAL)，、CAL高浓度 +3-MA组(加 5 mmol/L-3-甲基腺嘌呤)处理 48 h。采用细胞计数试剂盒( CCK-8)法检测细胞存活率；细胞自噬( MDC)染色检测观察自噬小体；实时荧光 PCR检测 Bcl-2相互作用蛋白 1(Beclin-1)、螯合体 1(P62)mRNA表达水平；流式细胞术检测各组细胞凋亡情况；蛋白质印迹法检测自噬及 PI3K/Akt/mTOR信号通路相关蛋白表达情况。结果与对照组细胞存活率、自噬水平、 Beclin-1，P62 mRNA，微管相关蛋白轻链 3BⅡ/Ⅰ(LC3BⅡ/LC3BⅠ)，Beclin-1与 P62蛋白、细胞凋亡率比较，高糖组细胞存活率［(100.07±4.81)%比(67.62±6.07)%］、自噬水平、 Beclin-1 mRNA和 LC3BⅡ/LC3B Ⅰ、Beclin-1蛋白表达降低， P62 mRNA和 P62蛋白表达、细胞凋亡率［( 2.86±0.47)%比( 15.42±1.78)%］增加( P<0.05)；与高糖组比较， CAL低、中、高浓度组细胞存活率、自噬水平、 Beclin-1 mRNA和 LC3BⅡ/LC3BⅠ、Beclin-1蛋白表达增加， P62 mRNA和 P62蛋白表达、细胞凋亡率降低( P<0.05)。与正常对照组细胞存活率、细胞凋亡率、磷酸化磷脂酰肌醇 3-激酶( p-PI3K)/PI3K、磷酸化蛋白激酶 B(p-Akt)/Akt、磷酸化哺乳动物雷帕霉素靶蛋白( p-mTOR)/mTOR蛋白表达比较，高糖模型组和 CAL高浓度 +3-MA组细胞存活率显著下降， p-PI3K/PI3K、p-Akt/Akt、p-mTOR/mTOR的蛋白表达水平显著升高，高糖模型组、 CAL高浓度组、 CAL高浓度 +3-MA组细胞凋亡率升高(P<0.05)；与高糖模型组比较， CAL高浓度组细胞存活率显著提升， p-PI3K/PI3K、 p-Akt/Akt、p-mTOR/mTOR蛋白表达显著降低， CAL高浓度组和 CAL高浓度 +3-MA组细胞凋亡率降低( P<0.05)；与 CAL高浓度组比较， CAL高浓度 +3-MA组细胞凋亡率升高( P<0.05)。结论毛蕊异黄酮可以通过抑制 PI3K/Akt/mTOR信号通路，促进自噬，保护 ARPE-19细胞免受高糖损害。 |
| 英文摘要: |
| Objective To investigate the effect of calycosin (CAL) on autophagy of human retinal pigment epithelial cells (ARPE-19) under high glucose through phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin (PI3K/Akt/mTOR) signaling pathway.Methods ARPE-19 cells were cultured in vitro from December 2022 to December 2023 and divided into control group (5.5mmol/L glucose), a hypertonic group (50 mmol/L mannitol), a high glucose group (50 mmol/L glucose), and low, medium and high con-centration of CAL groups (50 mmol/L glucose+25, 50, 75 μmol/L-CAL). After screening and determining the optimal concentration ofCAL (75 μmol/L), the cells were treated for 48 h and divided into a normal control group, a high glucose model group, a CAL high-con-centration group (50 mmol/L glucose + 75 μmol/L CAL), and a CAL high-concentration + 3-MA group (with additional 5 mmol/L 3-methyladenine). The cell survival rate was detected by Cell Counting Kit (CCK-8) method; autophagy (MDC) staining was used to ob-serve autophagosomes; the mRNA expression levels of Bcl-2 interacting protein 1 (Beclin-1) and sequestosome 1 (P62) were detected by real-time PCR; Apoptosis was detected by flow cytometry; Western blotting was used to detect the expression of autophagy and PI3K/Akt/mTOR signaling pathway related proteins.Results Compared with the control group, the cell survival rate, autophagy level, Be-clin-1, P62 mRNA, Microtubule associated protein light chain 3BⅡ/I (LC3BⅡ/LC3BⅠ), Beclin-1 and P62 protein, and apoptosis rate, while the cell survival rate [(100.07±4.81)% vs. (67.62±6.07)%], autophagy level, Beclin-1 mRNA, LC3BⅡ/LC3BⅠ, Beclin-1 protein expression decreased, P62 mRNA and P62 protein expression, and apoptosis rate [(2.86±0.47)% vs. (15.42±1.78)%] increased in the high glucose group (P>0.05). Compared with the high glucose group, the cell survival rate, autophagy level, Beclin-1 mRNA and LC3B Ⅱ/LC3BⅠ, Beclin-1 protein expression were increased, while P62 mRNA and P62 protein expression, apoptosis rate were decreasedin the low, medium, and high concentration groups of CAL (P<0.05). Compared with the normal control group, the cell survival rate, apoptosis rate, and protein expression of p-PI3K/PI3K, p-Akt/Akt, and p-mTOR/mTOR were significantly decreased in the high glucose model group and the high concentration of CAL + 3-MA group, while the protein expression levels of p-PI3K/PI3K, p-Akt/Akt, and p-mTOR/mTOR were significantly increased, and the cell apoptosis rates were significantly increased in the high model glucose group,the high concentration of CAL group, and the high concentration of CAL + 3-MA group, respectively (P<0.05). Compared with the highglucose model group, the cell survival rate of the high concentration of CAL group was significantly increased, the protein expression ofp-PI3K/PI3K, p-Akt/Akt, and p-mTOR/mTOR was significantly decreased, and the apoptosis rate of the high concentration of CALgroup and the high concentration of CAL + 3-MA group was decreased (P<0.05). Compared with the high concentration of CAL group, the apoptosis rate in the high concentration of CAL + 3-MA group was increased (P<0.05).Conclusion Calycosin can promote autoph-agy and protect ARPE-19 cells from high glucose damage by inhibiting PI3K/Akt/mTOR signaling pathway. |
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